A cellular hemoglobin-based oxygen carriers show promise for use as blood substitutes. They are basically prepared by extracting hemoglobin from red blood cells, purifying it to a very high degree, intramolecularly crosslinking it to ensure that it maintains its basic tetrameric 64 kilodalton molecular size, optionally oligomerizing it to a higher molecular weight, and preparing an aqueous solution thereof at appropriate concentration. Prior art references disclosing such blood substitutes include U.S. Pat. No. 4,857,636 Hsia, U.S. Pat. No. 5,250,665 Kluger et. al, a book entitled "Hemoglobin-based Red Cell Blood Substitutes" edited by R. M. Winslow, John Hopkins University Press (1992), and a book entitled "Blood Substitutes and Oxygen Carriers", edited by Thomas M. S. Chang, Marcel Dekker, Publisher, New York (1992).
One of the primary fields in which such blood substitutes are required for use is in association with surgical procedures. The reintroduction of oxygen into an ischemic tissue or organ can cause ischemia-reperfusion injury. Such injuries are generally accepted to be associated with the presence of oxygen-derived free radicals. A desirable oxygen carrier is one that delivers oxygen effectively, and at the same time prevents the increase in the oxygen-derived free radicals. It is suspected that hemoglobin-based solutions when used in biological systems may lead to an increase in oxygen free radicals, and provide catalytic heme iron.
There are several mechanisms by which the generation of oxygen-derived free radicals can occur. In one mechanism, in the presence of superoxide and hydrogen peroxide, heme iron (bound and/or released) can drive fenton reactions generating highly reactive perferryl or hydroxyl reactions. In a second mechanism, heme groups can auto-oxidize and release superoxide. In a third mechanism, modified hemoglobins may stimulate phagocyte respiratory burst activity and increase arachidonic acid metabolism, leading to an increase in superoxide production.
Endogenous antioxidant enzymes, namely superoxide dismutase (SOD) and catalase, catalyze the breakdown of superoxide and hydrogen peroxide respectively. Although not all reported results are consistent, many reports have shown that SOD and/or catalase are effective in reducing reperfusion injury and other free radical-mediated injury processes.
It is an object of the present invention to provide a novel crosslinked hemoglobin based oxygen carrier.
It is a further object to provide such a novel carrier which utilizes the beneficial properties of at least one endogenous enzyme.